GPR30 Deficiency Causes Increased Bone Mass, Mineralization, and Growth Plate Proliferative Activity in Male Mice

Estrogen regulation of the male skeleton was first clearly demonstrated in patients with aromatase deficiency or a mutation in the ERα gene. Estrogen action on the skeleton is thought to occur mainly through the action of the nuclear receptors ERα and ERβ. Recently, in vitro studies have shown that the G protein–coupled receptor GPR30 is a functional estrogen receptor (ER). GPR30-deficient mouse models have been generated to study the in vivo function of this protein; however, its in vivo role in the male skeleton remains underexplored. We have characterized size, body composition, and bone mass in adult male Gpr30 knockout (KO) mice and their wild-type (WT) littermates. Gpr30 KO mice weighed more and had greater nasal-anal length (p < .001). Both lean mass and percent body fat were increased in the KO mice. Femur length was greater in Gpr30 KO mice, as was whole-body, spine, and femoral areal bone mineral density (p < .01). Gpr30 KO mice showed increased trabecular bone volume (p < .01) and cortical thickness (p < .001). Mineralized surface was increased in Gpr30 KO mice (p < .05). Bromodeoxyuridine (BrdU) labeling showed greater proliferation in the growth plate of Gpr30 KO mice (p < .05). Under osteogenic culture conditions, Gpr30 KO femoral bone marrow cells produced fewer alkaline phosphatase–positive colonies in early differentiating osteoblast cultures but showed increased mineralized nodule deposition in mature osteoblast cultures. Serum insulin-like growth factor 1 (IGF-1) levels were not different. These data suggest that in male mice, GPR30 action contributes to regulation of bone mass, size, and microarchitecture by a mechanism that does not require changes in circulating IGF-1. © 2011 American Society for Bone and Mineral Research

Relative roles of TGF-β1 and Wnt in the systemic regulation and aging of satellite cell responses

Muscle stem (satellite) cells are relatively resistant to cell-autonomous aging. Instead, their endogenous signaling profile and regenerative capacity is strongly influenced by the aged P-Smad3, differentiated niche, and by the aged circulation. With respect to muscle fibers, we previously established that a shift from active Notch to excessive transforming growth factor-beta (TGF-β) induces CDK inhibitors in satellite cells, thereby interfering with productive myogenic responses. In contrast, the systemic inhibitor of muscle repair, elevated in old sera, was suggested to be Wnt. Here, we examined the age-dependent myogenic activity of sera TGF-β1, and its potential cross-talk with systemic Wnt. We found that sera TGF-β1 becomes elevated within aged humans and mice, while systemic Wnt remained undetectable in these species. Wnt also failed to inhibit satellite cell myogenicity, while TGF-β1 suppressed regenerative potential in a biphasic fashion. Intriguingly, young levels of TGF-β1 were inhibitory and young sera suppressed myogenesis if TGF-β1 was activated. Our data suggest that platelet-derived sera TGF-β1 levels, or endocrine TGF-β1 levels, do not explain the age-dependent inhibition of muscle regeneration by this cytokine. In vivo, TGF-β neutralizing antibody, or a soluble decoy, failed to reduce systemic TGF-β1 and rescue myogenesis in old mice. However, muscle regeneration was improved by the systemic delivery of a TGF-β receptor kinase inhibitor, which attenuated TGF-β signaling in skeletal muscle. Summarily, these findings argue against the endocrine path of a TGF-β1-dependent block on muscle regeneration, identify physiological modalities of age-imposed changes in TGF-β1, and introduce new therapeutic strategies for the broad restoration of aged organ repair

Estrogen Regulates the Satellite Cell Compartment in Females

Skeletal muscle mass, strength, and regenerative capacity decline with age, with many measures showing a greater deterioration in females around the time estrogen levels decrease at menopause. Here, we show that estrogen deficiency severely compromises the maintenance of muscle stem cells (i.e., satellite cells) as well as impairs self-renewal and differentiation into muscle fibers. Mechanistically, by hormone replacement, use of a selective estrogen-receptor modulator (bazedoxifene), and conditional estrogen receptor knockout, we implicate 17β-estradiol and satellite cell expression of estrogen receptor α and show that estrogen signaling through this receptor is necessary to prevent apoptosis of satellite cells. Early data from a biopsy study of women who transitioned from peri- to post-menopause are consistent with the loss of satellite cells coincident with the decline in estradiol in humans. Together, these results demonstrate an important role for estrogen in satellite cell maintenance and muscle regeneration in females

Enobosarm (GTx-024) modulates adult skeletal muscle mass independently of the androgen receptor in the satellite cell lineage

Androgens increase skeletal muscle mass, but their clinical use is hampered by lack of tissue selectivity and subsequent side-effects. Selective androgen receptor modulators (SARMs) elicit muscle-anabolic effects while only sparingly affecting reproductive tissues. The SARM GTx-024 (enobosarm) is being investigated for cancer cachexia, sarcopenia, and muscle wasting diseases. Here, we investigate the role of muscle androgen receptor (AR) in the anabolic effect of GTx-024. In mice lacking AR in the satellite cell lineage (satARKO), the weight of the androgen-sensitive levator ani muscle was lower, but decreased further upon orchidectomy. GTx-024 was as effective as dihydrotestosterone (DHT) in restoring levator ani weights to sham levels. Expression of the muscle-specific androgen-responsive genes S-adenosylmethionine decarboxylase and myostatin was decreased by orchidectomy and restored by GTx-024 and DHT in control mice, while expression was low and unaffected by androgen status in satARKO. In contrast, insulin-like growth factor IEa expression was not different between satARKO and control muscle, decreased upon castration, and was restored by DHT and GTx-024 in both genotypes. These data indicate that GTx-024 does not selectively modulate AR in the satellite cell lineage and that cells outside this lineage remain androgen-responsive in satARKO muscle. Indeed, residual AR positive cells were present in satARKO muscle, coexpressing the fibroblast-lineage marker vimentin. AR positive, muscle-resident fibroblasts could therefore be involved in the indirect effects of androgens on muscle. In conclusion, both DHT and GTx-024 target AR pathways in the satellite cell lineage, but cells outside this lineage also contribute to the anabolic effects of androgens.status: publishe

Insulin-like growth factor - Oestradiol crosstalk and mammary gland tumourigenesis

Development and differentiation of the mammary gland are dependent on the appropriate temporal expression of both systemically acting hormones and locally produced growth factors. A large body of evidence suggests that molecular crosstalk between these hormonal and growth factor axes is crucial for appropriate cell and tissue function. Two of the most important trophic factors involved in this process are the oestrogen (E) and insulin-like growth factor (IGF) molecular axes. The reciprocal crosstalk that exists between these pathways occurs at transcriptional/post-transcriptional and translational/post-translational levels regulate the expression and activity of genes involved in this process. In a clinical context an important consequence of such crosstalk in the mammary gland is the role which it may play in the aetiology, maintenance and development of breast tumours. Although oestradiol (E2) acting through oestrogen receptors α and β (ERα/β) is important for normal mammary gland function it can also provide a mitogenic drive to ER+ breast tumours. Therefore over several years anti-oestrogen therapeutic regimens in the form of selective oestrogen receptor modulators (SERMs - e.g. tamoxifen), aromatase inhibitors (AI e.g. anastrozole) or selective oestrogen receptor down regulators (SERDs - e.g. fulvestrant) have been used in an adjuvant setting to control tumour growth. Although initial response is usually encouraging, large cohorts of patients eventually develop resistance to these treatments leading to tumour recurrence and poor prognosis. There are potentially many routes by which breast cancer (BC) cells could escape anti-oestrogen based therapeutic strategies and one of the most studied is the possible growth factor mediated activation of ER(s). Because of this, growth factor modulation of ER activity has been an intensively studied route of molecular crosstalk in the mammary gland. The insulin-like growth factors (IGF-1 and -2) are amongst the most potent mitogens for mammary epithelial cells and there is accumulating evidence that they interact with the E2 axis to regulate mitogenesis, apoptosis, adhesion, migration and differentiation of mammary epithelial cells. Such interactions are bi-directional and E2 has been shown to regulate the expression and activity of IGF axis genes with the general effect of sensitising breast epithelial cells to the actions of IGFs and insulin. In this short review we discuss the evidence for the involvement of crosstalk between the insulin-like growth factor (IGF) and oestrogen axes in the mammary gland and comment on the relevance of such studies in the aetiology and treatment of BC